Use of affinity tags for purification
The use of affinity tags with the desired protein is a powerful approach for recombinant protein recovery, but the tag needs to be removed from the final product. Several affinity tags have been described for recombinant protein purification (Terpe, 2003). N-terminal hexa histidine tagged ricin B (HIS-RTB), the lactin subunit of ricin, was expressed in tobacco. The recombinant HIS-RTB was purified from tobacco leaves using lactose affinity column, and was found to be functional (Reed et al., 2005). His-tagged amarantin was expressed in tobacco and the presence of His tag was used for single-step purification of recombinant protein using immobilized metal-ion affinity chromatography (Valdez-Ortiz et al., 2005). The effects of three affinity tags, i.e. eight amino acid tag StrepII, His6 and 181-amino acid Tandem Affinity Purification (TAP) tag were studied for the purification of recombinant membrane anchored protein kinase. The protein purified using His6 tag was of low purity whereas the recombinant proteins having TAP or StrepII tag were purified to homogeneity. While StrepII-tag purification achieved high yield and purity which was comparable to that obtained with TAP-tag, it was considerably easier and faster (Witte etal.,2004). In addition to aid in the purification of recombinant proteinfrom plants (Conley et al., 2009; Joensuu et al., 2009), elastin like polypeptide tag has also been found to enhance expression of the protein target (Conley et al., 2009). The possibilities of using histamine, tryptamine, phenylamine or tryamine as affinity tags for affinity purification of recombinant proteins have also been evaluated (Platis and Labrou, 2008). The protein splicing elements (inteins) that can catalyse self-cleavage (Liu, 2000) have also been utilized in protein purification purposes. The SMAP 29, a mammalian antimicrobial peptide (Bagella et al., 1995), was expressed with intein fusion tag in tobacco plants.
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